Dysbiosis and Rebiosis of the Periodontal Microbiome

The human body supports the growth of a wide array of microbial communities in the gastro-intestinal and urogenital tracts and on the surface of the skin. Together, these communities of bacteria are referred to as the human microbiome. It is widely acknowledged that the human microbiome plays a significant role in human biology through its influence on human development, physiology, immunity and nutrition. Although the composition of the human microbiome has received considerable attention in recent years, the precise mechanisms whereby the microbial communities mediate disease and maintain health remain uncertain. However, recent studies have shown that several chronic diseases of the mouth and gastro-intestinal tract are associated with alterations to the composition of the entire microbiome. This is referred to as dysbiosis where there are major and harmful shifts in the relative abundancies of individual components of the microbiome compared to the abundancies found in health.

Periodontal disease is an example of these conditions where dysbiosis of the normal microbiology takes place. This disease is one of the most common inflammatory diseases of humans leading to tooth loss in approximately 20% of the population and a significant cost to the NHS. It is also thought to be a risk factor for the development of other diseases including cardiovascular disease and type II diabetes. Current treatment methods involve thorough cleaning of the tooth surfaces below the gum margin, sometimes supplemented by antibiotic therapy. However, even with regular treatment episodes this is not always successful. There is, therefore, an urgent unmet clinical need to develop newer methods of both diagnosis, prevention and treatment of this condition. Given the current concerns about the development of resistance to antibiotics, newer treatments for periodontal disease should aim to avoid the use of these valuable agents for more acute and life threatening conditions.

Attempts to study these processes in humans have had limited success to date - possibly because of the large variation in clinical disease in human populations and because of the need for longitudinal studies over a long time scale: disease in humans is a slowly progressive disease. We have shown in studies using mice that introduction of the human periodontal organism, Porphyromonas gingivalis, into the mouth causes dysbiosis of the oral microbiome and the development of periodontal disease. P. gingivalis causes these major shifts to the normal microbiology even though it is present in very low quantities. We therefore refer to this bacterium as a keystone pathogen able to manipulate the composition of the normal bacteria in the mouth even though it is present in low abundance. Similar keystone pathogens have now been described in other inflammatory diseases of humans. We have also shown that the P. gingivalis disease- associated microbiome is very stable and can be transferred into healthy recipient mice and cause disease. This system therefore provides an ideal experimental model to determine mechanisms of dysbiosis of the oral microbiome and how this may be reversed in order to restore health.

Specifically, in this investigation, we aim:
1. To determine how a keystone pathogen - in this case P. gingivalis - causes microbial dysbiosis
2. To establish what are the functional properties of a dysbiotic periodontal microbiome compared to the microbiome in health
3. To determine which component(s) or properties of a normal symbiotic microbiome may be used to reverse dysbiosis and hence restore health in a diseased individual

The overall aim of these investigations is to form a basic understanding of the mechanisms of dysbiosis of the periodontal microbiome and its potential reversal to a symbiotic state (rebiosis). Through these experiments we aim to provide the basis for the development of novel approaches to the treatment and prevention of human disease.

Dysbiosis of the oral microbiome in periodontal disease is a hallmark of this condition. Understanding the mechanism of dysbiosis, its functional relevance to disease and strategies to achieve reversal of dysbiosis to restore health have been hindered by the lack of a suitable animal model. Our recent investigations using the mouse model of this disease have demonstrated that the human periodontal bacterium Porphyromonas gingivalis acts as a keystone pathogen in manipulating the normal commensal microbiome into a dysbiotic condition even when present at low abundance. Furthermore we have also shown that this dysbiotic microbiome is causative of disease rather than a consequence of the altered environment in this inflammatory condition. We aim to build upon these finding using this tractable model system in this application.

The specific aims are:
1. Determine the mechanism of P. gingivalis induced dysbiosis of the periodontal microbiome by assessment of the dysbiotic potential of P. gingivalis genetic mutants in candidate genes with particular reference to the role of the Type IX secretion system and the production of outer membrane vesicles
2. Functional characterisation of the dysbiotic periodontal microbiome compared to the symbiotic microbiome characteristic of health through metagenomic and meta-transcriptomic analyses
3. Reversal of dysbiosis: Rebiosis of the periodontal microbiome. Informed by the findings of the preceding aims we will identify organisms or consortia of organisms whose presence/ transcriptional activities are consistent with symbiosis and periodontal health and apply these in bacterial replacement investigations in order to reverse dysbiosis and restore health in this model system.

These investigations will form the basis for the design of future translational investigations in humans with the overall aim of the development of novel treatment approaches to periodontal disease.

Study of the human oral microbiome is one of the most advanced in this field in terms of the understanding of the number and types of different species which inhabit this niche in both health and disease. However, the interpretation of this information is hindered by the lack of well characterised animal models to enable identification of those bacterial species, or consortia of species, which are functionally important in the maintenance of health and the development of disease. This is particularly important in the case of periodontal disease, where it is well recognised that there is a dramatic shift in the composition of the microbiome. The standard treatment of periodontal disease, namely debridement of tooth surfaces with/without supplemental antibiotic therapy has not changed for decades. This is both time-consuming, a significant use of valuable antibiotics in the community and in some cases of limited value to prevention of the progression of disease. New approaches are therefore a priority.

In this application, we aim to characterise the mechanisms of dysbiosis and rebiosis of the oral microbiome of the mouse in order to study the influence of the microbiome and its dysbiosis on the development of periodontal disease in a genetically well characterised host background. The results of the mouse microbiome analysis will be of benefit to the international periodontal research community, many of whom use the mouse model of periodontal disease as part of the research investigations, and we will therefore make the 16S rRNA data and transcriptome data publically available. Moreover, a potential outcome of these investigations will be the identification of bacterial species which have a protective effect on the oral tissues, analogous to the properties of, for example, Faecalibacterium prauznitzii and polysaccharide A of Bacteroides fragilis in the intestine, and hence future developments may lead to the development of novel therapeutic approaches in collaboration with other academic groups and industry involving bacterial replacement therapy or a probiotic approach.